Amitriptyline is a potent antagonist to histamine H1 and H2 receptors. High affinity binding of 3H-amitriptyline can be used to specifically label the H1 receptor as shown by KD values of H1 antagonists that agree with those of known H1 systems. These methods are also being adapted to study amitriptyline binding to H2 receptors. The H2 receptor density (Bmax) will be measured in subcellular and regional fractions of guinea pig, rat and mouse brain. In homogenates the H2 but not the H1 receptor is linked to adenylate cyclase; in brain slices both receptors have been implicated. To test the latter we will study the effect of selective H1 and H2 agonists and antagonists on the histamine activation of cyclic AMP synthesis in brain slices and measure the KD values for the antagonists, the definitive way of identifying the receptors. Tricyclic antidepressants are H1 and H2 antagonists but their mode of receptor antagonism may be different from those of the conventional ones, e.g., mepyramine and cimetidine. KD values for some of these drugs will be obtained by studing their competition with both 3H-mepyramine and 3H-amitriptyline to the H1 receptor. Since there is no re-uptake of histamine, its enzymatic methylation may serve as the inactivation mechanism. This will be tested with inhibitors of histamine methyltransferase (HMT) which should potentiate the activation of adenylate cyclase by histamine. Since all evidence indicates that formation of methylhistamine in brain depends on histamine release, mythylhistamine content may reflect the functioning of histaminergic nerves. The action of some drugs known to affect this system will be studied on the regional content of histamine and methylhistamine, the latter measured by a method recently developed by us.